The present invention relates to methods for the modulation of nuclear receptor mediated processes, compounds useful therefor and methods for the identification and use of such compounds.
The actions of steroids, retinoids and thyroid hormones are mediated by intracellular nuclear receptors whose coordinate activity defines the physiological response (Mangelsdorf and Evans, Cell 83:841-850 (1995)). These receptors are all structurally related and constitute a superfamily of nuclear regulatory proteins that modulate gene expression in a ligand-dependent fashion. Previous studies have demonstrated that the 9-cis retinoic acid receptor (RXR) serves as a common heterodimeric partner for thyroid hormone receptor (TR), retinoic acid receptor (RAR), vitamin D receptor (VDR), prostanoids (PPAR), as well as numerous orphan receptors (Kliewer et al. (1992) Nature 355:446-449).
Transcriptional repression is an intrinsic part of endocrine physiology and contributes to feedback regulation associated with the inhibition of the physiologic response. Indeed, the thyroid hormone receptor is converted to an oncogene by mutations which block hormone binding and create a constitutive transcriptional repressor (Damm et al. EMBO J. 6:375-382 (1987), Nature 3:593-597 (1989); Graf and Beug Cell 34:7-9 (1983); Sap et al. Nature 340:242-244 (1989)). Multiple studies on transcriptional silencing by verbA and the non-liganded thyroid hormone receptor suggest that repression is required for oncogenesis and that this process is mediated by a diffusible co-factor(s) that associates with the ligand binding domain (LBD) (Baniahmad et al. Mol. Cell. Biol. 15:76-86 (1995); Casanova et al. Mol. Cell. Biol. 14:1756-1765 (1994)).
Transcriptional co-repressors (SMRT and N-CoR) have recently been identified that associate with non-liganded receptors resulting in suppression of basal transcriptional activity (see, for example, Chen and Evans Nature 377:454-457 (1995); Chen et al. PNAS 93:7567-7571 (1996); Horlein et al. Nature 377:397-404 (1995); and Sande and Privalsky Mol. Endo. 10:813-825 (1996)).
While the mechanism of this repression is not known, chromatin remodeling has been suggested to be a component of transcriptional regulation (for review see Wolffe and Pruss. Curr. Biol. 6:234-237 (1996): Felsenfeld Cell 86:13-19 (1996)). Indeed, it has been suggested that specific transcriptional activation may be involved in local changes in chromatin structure. In fact, it has recently been demonstrated that nuclear hormone receptors may utilize the CREB binding protein (CBP) or its homolog p300 (Janknecht and Hunter Nature 383:22-23 (1996)), to function as a nuclear receptor co-factor (Chakravarti, et al. Nature 383:99-103 (1996); Hanstein et al. PNAS 93:11540-11545 (1996); Kamei et al. Cell 85:403-414 (1996); Yao et al. PNAS 93:10626-10631 (1996)). In addition to CBP/p300, multiple hormone-dependent and independent associated co-factors have been characterized (Fondell et al. PNAS 93:8392-8333 (1996)).
Of particular interest is the recent demonstration that CPB/p300 associates with the histone acetylase P/CAF (Yang X-J et al. Nature 382:319-324 (1996)) which displays significant sequence homology to the yeast transcription activator GCN5, also known to be a histone acetylase (Brownell et al. Cell 84:843-851 (1996)). Further, CBP/p300 harbors intrinsic histone acetyltransferase activity, resulting in alternative or perhaps simultaneous histone acetylation (Ogryzko et al. Cell 87:953-959 (1996)). The notion that multiple transcriptional co-activators possess acetylase activity suggests that their recruitment to a DNA template would locally destabilize nucleosomes, creating a permissive state for promoter activation.
The symptoms of certain neoplastic diseases have been ameliorated by the administration of retinoids. However, some patients afflicted with acute promyelocytic leukemia (APL) respond poorly (if at all), to retinoid differentiation therapy, while others enter disease remission following high doses of retinoic acid treatment. Although APL cell lines have been identified as being retinoid sensitive, there has been no evidence to explain the distinct retinoid responses in APL patients.
Accordingly, there is a need in the art for a further understanding of the interaction(s) between the various components involved in regulation of hormone mediated processes. A clearer understanding of these processes will facilitate the development of methods to modulate hormone mediated processes, as well as assays for the identification of compounds useful for such modulation. Moreover, there is a need in the art for novel approaches for treating human cancer. These and other needs in the art are addressed by the present invention.
In accordance with the present invention, we have discovered that histone deacetylase associates with hormone receptor complexes and contributes to the repression thereof. We have further discovered that exposure of a repressed system to histone deacetylase inhibitors relieves this repression. Thus, histone deacetylase inhibitors have been found to be useful for the activation of genes responsive to hormone receptors.
In accordance with another aspect of the invention, formulations useful for modulation of hormone-mediated processes have been developed. In addition, assays have been developed for the identification of compounds useful to modulate the above-described processes. In accordance with another aspect, methods of employing such compounds to modulate or enhance hormone mediated processes, such as human cancers.
In accordance with the present invention, SMRT and components of the histone deacetylase complex have been discovered to function as effectors of cellular transformation. Specifically, the interaction of SMRT with PLZF-RARxcex1 and PML-RARxcex1 has been discoved to play a critical role in the pathogenesis of acute promyelocytic leukemia (APL). These findings provide the first direct link between nuclear receptor cofactors and human cancer by providing evidence that a transcriptional corepressor may directly contribute to human cancers. Moreover, pharmacological manipulation of the association/dissociation of nuclear receptor cofactors will prove to be beneficial as it provides novel approaches to treatment of human diseases.